Development of a novel crAss-like phage detection method with a broad spectrum for microbial source tracking

Area of Science:

• Microbiology
• Virology
• Environmental Science

Background:

• CrAssphage is a key human gut virus used for microbial source tracking (MST) of fecal contamination.
• Recent discoveries of crAss-like phages (CLPs) have led to the new viral order Crassvirales.
• CLPs offer potential for expanded MST applications beyond traditional crAssphage detection.

Purpose of the Study:

• To investigate the prevalence of CLPs in South Korea.
• To develop a CLP detection system for MST.
• To identify novel, human-specific MST markers.

Main Methods:

• Metagenomic analysis of human fecal samples to identify CLPs.
• Categorization of CLPs into genera based on major head protein (MHP) sequence similarity.
• Development and evaluation of PCR primers targeting MHP sequences.
• Testing CLP presence in human and animal fecal samples.

Main Results:

• Thirteen CLPs were identified and classified into seven genera.
• CLPs were detected in 91.52% of human fecal samples.
• Genus VI CLPs were most prevalent in humans (64.4%), exceeding p-crAssphage detection rates.
• CLPs were largely absent in animal feces, except for raccoons harboring specific genera.

Conclusions:

• Genus VI CLPs represent a highly effective and prevalent human-specific MST marker.
• The expanded CLP detection broadens the scope of viral MST for human fecal contamination.
• These findings can inform public health strategies and reduce costs associated with environmental monitoring.